In the present study we examined the hypothesis that fatty acid oxidation is less during high-intensity exercise than during moderate-intensity exercise because of inhibition of long-chain fatty acid entry into the mitochondria. Six volunteers exercised at 40% peak oxygen consumption for 60 min and at 80% for 30 min on two different occasions. [1-13C]oleate, a long-chain fatty acid, and [1-14C]octanoate, a medium-chain fatty acid, were infused for the duration of the studies. Lipids and heparin were infused during exercise at 80% to prevent the expected decrease in plasma free fatty acid (FFA) concentration. Plasma oleate and total FFA availability were similar in the two experiments. Our data suggest that, in addition to sub-optimal FFA availability, fatty acid oxidation is likely limited during high-intensity exercise because of direct inhibition of long-chain fatty acid entry into mitochondria.